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International audience

International audience

Multiple sequence alignment (MSA) is a prerequisite for most phylogenetic analyses. Aligning sequences to unravel residue homology is a challenging task that has been the focus of much attention in recent decades. Research in this field has been extremely active from both theoretical and practical standpoints. Numerous tools have been developed to align sequences and, more recently, to post-process those alignments and filter out their most dubious parts. Whether or not the inclusion of alignment filtering in a phylogenetic pipeline improves the quality of the inferred phylogenies is still debatable. The goal of this chapter is not to provide an exhaustive list of all tools available to produce or filter an MSA, but rather to cover the limitations of current alignment methods and their causes, to highlight key differences among MSA filtering methods and provide some practical MSA filtering guidelines. We consider that filtering methods can be subdivided into two main categories. The first one includes methods that filter MSA by entirely removing some sites or sequences from the MSA. The second category contains MSA filtering methods that mask residues and are able to extract some pieces of information from a site or sequence, while disregarding the remaining information-we called these picky-filtering methods. In our benchmark, the filtering methods that perform best are, as expected, in the picky category. When inferring phylogenies, MSA filtering impacts the inferred tree topology but it also seems to significantly improve branch length estimations, especially when a picky-filtering method is used. How to cite: Vincent Ranwez and Nathalie Chantret (2020).

Native African cereals (sorghum, millets) ensure food security to millions of low-income people from low fertility and drought-prone regions of Africa and Asia. In spite of their agronomical importance, the genetic bases of their adaptations to natural and anthropogenic selective pressures are still not well understood. Here we focus on Sorghum bicolor, which is the fifth cereal worldwide for grain production and constitutes the staple food for around 500 million people. The release of the genome sequence, its phylogenetic proximity with other important C4 species (maize, switchgrass, sugarcane) and its low genome complexity contribute to the interest on S. bicolor on a more fundamental level. We leverage transcriptomic and genomic resources to address the adaptive consequences of the domestication process. Despite gene flow with the wild pool, a clear domestication syndrome is visible in cultivated sorghum (plant architecture, shattering, dormancy and seed weight). Gene expression and nucleotide variability were analyzed in 11 cultivated and 9 wild accessions. The cultivated compartment harbors 30% lower genetic diversity than the wild pool, which supports the occurrence of a genetic bottleneck in the domestication history of sorghum. Furthermore, the crop gene pool showed polymorphism patterns in coding regions indicative of lower efficacy of purifying selection, suggesting that sorghum suffers from the cost of domestication. 949 genes are significantly differentially expressed between the two compartments. Among them, genes involved in internode elongation and cytoskeleton organization are over-expressed, while genes linked to photosynthesis and reduction-oxidation processing of seed storage proteins are under-expressed in the cultivated accessions. Further analyses revealed that domestication reduced significantly the variability of the isoform expression balance (expression ratio of the alternative proteins coded by a single gene) in cultivated sorghum. Overall, these analyses pave the way towards the identification of key domestication genes valuable for genetic resources characterization and breeding purposes.

Oaks are an important part of our natural and cultural heritage. Not only are they ubiquitous in our most common landscapes1 but they have also supplied human societies with invaluable services, including food and shelter, since prehistoric times2. With 450 species spread throughout Asia, Europe and America3, oaks constitute a critical global renewable resource. The longevity of oaks (several hundred years) probably underlies their emblematic cultural and historical importance. Such long-lived sessile organisms must persist in the face of a wide range of abiotic and biotic threats over their lifespans. We investigated the genomic features associated with such a long lifespan by sequencing, assembling and annotating the oak genome. We then used the growing number of whole-genome sequences for plants (including tree and herbaceous species) to investigate the parallel evolution of genomic characteristics potentially underpinning tree longevity. A further consequence of the long lifespan of trees is their accumulation of somatic mutations during mitotic divisions of stem cells present in the shoot apical meristems. Empirical4 and modelling5 approaches have shown that intra-organismal genetic heterogeneity can be selected for6 and provides direct fitness benefits in the arms race with short-lived pests and pathogens through a patchwork of intra-organismal phenotypes7. However, there is no clear proof that large-statured trees consist of a genetic mosaic of clonally distinct cell lineages within and between branches. Through this case study of oak, we demonstrate the accumulation and transmission of somatic mutations and the expansion of disease-resistance gene families in trees.

Poster presented at EUCARPIA Genetic Resources 2017 in MontpellierUnderstanding the genetic architecture related to plant adaptation and environmental heterogeneity is a crucial issue due to the current context of climate change. Local climatic conditions likely constitute an important selective pressure on genes underlying important fitness-related traits like flowering time. To test this assumption and to identify flowering candidate genes involved in plant adaptation to climate variation, we analyse patterns of nucleotide diversity in 224 flowering time candidate genes on a large core-collection of Medicago truncatula, spanning different climatic gradients. We found that flowering time mediates adaptation to climate heterogeneity in M. truncatula, mainly by variation at genes located upstream the flowering pathways, close to the environmental stimuli. Variables related to the annual precipitation regime reflected selective constraints on flowering time genes better than the other variables tested (temperature, altitude, latitude or longitude). By comparing phenotype and climate associations, we identified 12 flowering genes as the most promising candidates to be involved in the phenological response to climate. Four of them were located in the known M. truncatula QTL region for flowering time in chromosome 7, which gives support to our findings. However, climate and flowering associations also highlighted largely distinct gene sets, suggesting different genetic architectures for adaptation to climate and flowering onset. This study illustrates the importance of the collection and management of PGR (including crop wild relatives).

Poster presented at the Society for Molecular Biologu and Evolution Meeting 2012 in DublinFlowering time is a phenotypic character potentially implied in the adaptive responses to different selective pressures like climate, pollinators, herbivores and pathogens. The Mediterranean basin, which is the area of distribution of the legume model Medicago truncatula, constitutes an ideal framework to address the effect of heterogeneous climatic conditions on adaptive characters. In this study, we examine the natural diversity of flowering genes in the annual autogamous M. truncatula and put it in relation with climatic variables and phenotypic characteristics. For this, we use next generation sequencing data for a sample of 192 accessions covering the whole distribution area of the species. First, we used a multivariate method (the Discriminant Analysis of Principal Components) and nucleotide polymorphism at 34,550 intergenic polymorphic sites distributed over the entire genome to check for the presence of neutral genetic structure within the sample. We found two major genetic groups, corresponding to western populations (from the Iberian Peninsula and Morocco) and eastern populations (from France and Algeria to the Middle East) respectively. Second, in each group, we applied different regression methods to identify sites potentially involved in the adaptation to climatic conditions. Preliminary results on a small subset of loci (330 SNP) point to 8-30% significant associations, depending on the method. These loci are potential candidates to be involved in local adaptation.

Sorghum bicolor is the fifth cereal worldwide for grain production, constituting the staple food for a round 500 million people. The release of its genome sequence, its phylogenetic proximity with other important C4 species (maize, switchgrass, sugarcane) and its low genome complexity contribute to its interest on a more fundamental level. Sorghum bicolor wild relatives (ssp. verticilliflorum) and domesticated (ssp. bicolor) pools are inter-fertile and intense gene flow occurs. However, a clear domestication syndrome is visible (plant architecture, shattering, dormancy and seed weight). ln spite of its agronomical Importance, the evolutionary dynamics of sorghum domestication and the genetic bases of adaptations to natural and anthropogenic pressures are not well understood. Here we leverage transcriptomic and genomic resources to improve our knowledge on the evolutionary history of S. bicolor, with a particular focus on the consequences of the domestication process. As a first step, gene expression and nucleotide variability of the expressed genes were analyzed in 11 cultivated and 9 wild accessions selected to maximize the genetic coverage of these two pools. The cultivated compartment harbors 30% lower genetic diversity than the wild pool, which supports the occurrence of a genetic bottleneck in the domestication history of sorghum. More than 900 genes are significantly differentially expressed between the two compartments. Among them, genes involved in photosynthesis and in reduction-oxidation processing of seed storage proteins are over-represented. Further analyses revealed that domestication reduced significantly the variability of the isoform expression balance (expression ratio of the alternative proteins coded by a single gene) in cultivated sorghum. Scans for local adaptation and positive selection on large cultivated and wild panels genotyped with genotyping by sequencing approaches are ongoing. Overall, these analyses pave the way towards the identification of key domestication genes that cou Id be useful for conservation strategies but also for breeding purposes.

National audience

AGAP : équipe ID / GE2pop; Leucine-Rich Repeats Receptor-Like Kinase (LRR-RLK) genes represent a large and complex gene family in plants, mainly involved in development and stress responses. These receptors are composed of an LRR-containing extracellular domain (ECD), a transmembrane domain(TM) and an intracellular kinase domain (KD). To provide new perspectives on functional analyses of these genes in model and non-model plant species, we performed a phylogeneticanalys is on 8,360 LRR-RLK receptors in 31 angiosperm genomes (8 monocots and 23dicots). We identified 101 orthologous groups (OGs) of genes being conserved among almost all monocot and dicot species analyzed. We observed that more than 10% of these OGs are absent in the Brassicaceae species studied. We show that the ECD structural features are not always conserved among orthologs, suggesting that functions may have diverged in some OG sets. Moreover, we looked at targets of positive selection footprints in 12 pairs of OGs and noticed that depending on the subgroups, positive selection occurred more frequently either in the ECDs or in the KDs.

AGAP : équipe GE2pop; Domestication is known to strongly reduce genomic diversity through population bottlenecks. The resulting loss of polymorphism has been thoroughly documented in numerous cultivated species. Here we investigate the impact of domestication on the diversity of alternative transcript expressions using RNAseq data obtained on cultivated and wild sorghum accessions (ten accessions for each pool). In that aim, we focus on genes expressing two isoforms in sorghum and estimate the ratio between expression levels of those isoforms in each accession. Noticeably, for a given gene, one isoform can either be overexpressed or underexpressed in some wild accessions, whereas in the cultivated accessions, the balance between the two isoforms of the same gene appears to be much more homogenous. Indeed, we observe in sorghum significantly more variation in isoform expression balance among wild accessions than among domesticated accessions. The possibility exists that the loss of nucleotide diversity due to domestication could affect regulatory elements, controlling transcription or degradation of these isoforms. Impact on the isoform expression balance is discussed. As far as we know, this is the first time that the impact of domestication on transcript isoform balance has been studied at the genomic scale. This could pave the way towards the identification of key domestication genes with finely tuned isoform expressions in domesticated accessions while being highly variable in their wild relatives.

AGAP : Equipe GE2pop; International audience; Common root rot, due to Aphanomyces euteiches, is a very damaging disease of legumes, especially pea, in many countries. Several consistent quantitative trait loci controlling partial resistance were identified in different pea genetic pools. Genomic basis of resistance to A. euteiches was investigated in the model legume Medicago truncatula in order to identify candidate genes underlying Aphanomyces resistance loci and analyze synteny of resistance loci between pea and M. truncatula. High density genome-wide association mapping was conducted in a collection of 179 M. truncatula accessions, for resistance to five reference A. euteiches isolates of main pea and alfalfa pathotypes. Two main genomic regions carrying candidate SNPs were associated with resistance. The first one corresponded to an F-box protein encoding gene at the prAe1/AER1 loci on chromosome 3 and the second one to a NBS-LRR resistance protein encoding gene underlying a new resistance locus on chromosome 4. The candidate genomic regions identified in M. truncatula were projected to a pea consensus genetic map carrying 1250 SNP markers anchored to M. truncatula genome. The F-box gene syntenic region co-located with the minor QTL Ae-Ps3.2 on pea LGIII and the NBS-LRR gene syntenic region was positioned about 50 cM far from the major QTL Ae-Ps7.6 on pea LGVII. Work is in progress to precise synteny between Aphanomyces resistance loci in pea and M. truncatula and to search for pea genes orthologous to the M. truncatula candidate genes identified

National audience

Background: Recurrent gene duplication and retention played an important role in angiosperm genome evolution. It has been hypothesized that these processes contribute significantly to plant adaptation but so far this hypothesis has not been tested at the genome scale.[br/] Results: We studied available sequenced angiosperm genomes to assess the frequency of positive selection footprints in lineage specific expanded (LSE) gene families compared to single-copy genes using a dN/dS-based test in a phylogenetic framework. We found 5.38% of alignments in LSE genes with codons under positive selection. In contrast, we found no evidence for codons under positive selection in the single-copy reference set. An analysis at the branch level shows that purifying selection acted more strongly on single-copy genes than on LSE gene clusters. Moreover we detect significantly more branches indicating evolution under positive selection and/or relaxed constraint in LSE genes than in single-copy genes.[br/] Conclusions: In this – to our knowledge –first genome-scale study we provide strong empirical support for the hypothesis that LSE genes fuel adaptation in angiosperms. Our conservative approach for detecting selection footprints as well as our results can be of interest for further studies on (plant) gene family evolution.

Communication orale; Le bassin mediterraneen, aire de distribution naturelle de la Legumineuse modele Medicago truncatula, constitue un cadre ideal pour etudier l’impact des conditions climatiques sur la variation des caracteres adaptatifs. La date de floraison est un caractere potentiellement implique dans la reponse a un ample spectre de pressions selectives : climat, herbivorie, pathogenes, pollinisateurs, et explique une bonne proportion de la variation de la distribution geographique des especes sauvages et cultivees. Dans cette etude nous examinons la diversite naturelle des genes impliques dans la variation de la date de floraison chez M. truncatula (Hecht et al. 2005), espece annuelle et autogame, pour la mettre en relation avec les variables climatiques et des caracteres de floraison . Pour cela, nous utilisons un jeu de donnees issu du sequencage haut debit de 192 plantes qui representent la totalite de l’aire de distribution de l’espece. Une methode multivariee, l'Analyse Discriminant des Composantes Principales (DaPC ; Jombart et al. 2010), a ete utilisee pour inferer la structure genetique neutre de cet echantillon. L’information de 34.550 sites nucleotidiques intergeniques repartis sur l'ensemble du genome montre une partition majeure en deux groupes geographiques : un groupe occidental regroupant des plantes originaires de la peninsule iberique et du Maroc et un deuxieme groupe plus oriental. La relation entre la variation genotypique et la variation environnementale a ete examinee a l'aide de methodes de regression, en tenant compte de la structure genetique et de l’apparentement entre individus intra groupe, pour reperer les locus potentiellement impliques dans la reponse aux conditions climatiques. Les resultats preliminaires sur un sous-echantillon de 330 locus SNP montrent jusqu'a 30% d'associations significatives. La validation de ces locus potentiellement impliques dans l'adaptation locale a l'aide de donnees phenotypiques, par genetique d’association, est en cours.

Whole genome duplications (WGDs) as well as small-scale duplications (SSDs) occurred frequently in plant evolution. In combination with a relatively high retention rate of novel genes, duplication events seem to be an important factor in plant genome evolution. Indeed, a large fraction of angiosperm diversity was suggested to stem from duplication followed by adaptive specialization of paralogous gene copies. Thanks to recent progress in genome sequencing, we can now address the question whether positive selection is a driving force in the evolution of duplicated genes. We searched a plant genome database for cases where recent successive duplication events followed by retention resulted in at least six gene copies. Those ultraparalogous gene clusters were tested for positive selection using codon substitution models implemented in PAML. We found that choosing the appropriate alignment and cleaning method is crucial as the codeml site-model is highly sensitive to alignment errors. Overall, 5-10% of all clusters showed evidence for positive selection. The banana genome shows very little evidence for selection at ultraparalogs, whereas duplicated genes in rice and Brachypodium seem to have undergone positive selection more frequently than the average. However, these results could also be explained by differences in genome coverage and quality.

National audience

L'analyse de la liaison statistique entre allèles à 2 locus, ou déséquilibre de liaison (DL), a été abordée simultanément chez le riz, le sorgho, le blé, l'hévéa et Medicago truncatula en utilisant des échantillons tirés de collections de ressources génétiques. Des stratégies utilisant des BACs ont permis de développer des marqueurs dans des zones localisées. L'utilisation des séquences terminales des BACs ciblés a été efficace pour définir des marqueurs microsatellites. Par ailleurs, des amorces ont été développées pour séquencer directement des fragments géniques. Le haut niveau de polymorphisme des marqueurs microsatellites a pu nécessiter des adaptations pour le calcul du DL. La structure génétique observée sur les plantes cultivées génère un fort bruit de fond de DL, tandis que dans les collections d'espèces sauvages autogames, le DL reste faible même à courte distance, décroissant rapidement au delà de 20 kb. A l'intérieur des groupes définis pour les espèces cultivées, le DL est plus faible et décroît assez rapidement, parfois de manière différente selon les groupes, de 100 kb environ pour le riz à 500 kb pour le sorgho. Une bonne connaissance de la diversité de l'espèce apparaît donc nécessaire avant de réaliser des études d'associations qui paraissent dès lors réalisables en utilisant les accessions contenues dans les collections de ressources génétiques.

Correspondance et tirés à part : tbata@daimi.au.dk; National audience; The comparative study of patterns of genetic diversity in crops and their wild ancestors coupled with recent advance in coalescent theory opens the way for making some historical inference on the demographic (and possibly selective) processes that have shaped genetic diversity in crops since domestication. Demographic events affect genome-wide levels of polymorphisms and the spectrum of allelic frequencies. Some forms of selection, in particular recent selective sweeps are expected to reduce diversity locally that is at the target of selection and in surrounding areas in the genome. Although a full reconstruction of the demographic history seems impossible based on patterns of polymorphism alone, contemporary polymorphism data can be used to calibrate simplified demographic models that can account for amounts and patterns of currently observable diversity. We illustrate that approach in two crop species with contrasted histories and mating systems. First, patterns of diversity are studied in the (allo)tetraploid wheat (Triticum turgidum) complex a species with a mixed mating system dominated by high levels of selfing. The intensity of bottlenecks associated with major historical episodes of domestication and subsequent selection are quantified using microsatellite markers. Patterns of microsatellite diversity are also compared with patterns of sequence polymorphism detected at the Gsp-1 locus. We find that domestication and subsequent selection resulted in a massive loss of genetic diversity and estimated a several fold reduction in the long term effective size realized in cultivated wheats relative to their wild ancestors (even after accounting for the probable loss of diversity in wild populations due to extensive habitat loss since Neolithic times). Next we analyze patterns of nucleotide diversity in alfalfa (Medicago sativa) an autotetraploid and outcrossing crop species. Sequence polymorphism among individuals spanning a wide geographic distribution was surveyed and analyzed at two intronic regions. These were located in respectively the alfalfa NADHdependent glutamate synthase and PO146, a pollen pectate lyase-like gene. These genes were chosen while searching in GenBank a public genetic sequence database for nucleotidic sequences of M. sativa and not chosen with any phenotypic à priori. This allowed us to i) quantify the extent of genetic divergence between wild and cultivated populations and thereby assess the impact of domestication on this divergence, ii) assess the originality of the genetic makeup of wild Spanish populations of alfalfa relative to the rest of extant population throughout the species range, iii) determine whether there is likely ongoing (recent) gene-flow among wild and cultivated populations in Spain

*INRA, Station d'Amélioration des Plantes, 35653 Le Rheu cedex Diffusion du document : INRA, Station d'Amélioration des Plantes, 35653 Le Rheu cedex Diplôme : Dr. Ing.

*INRA, Centre de Rennes Diffusion du document : INRA, Centre de Rennes Diplôme : DEA-DAA